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Two-channel image analysis method for the screening of OBOC libraries

Helmer, Dorothea 1; Brahm, Kevin; Helmer, Christian 2; Wack, Julia Susanne; Brenner-Weiss, Gerald 3; Schmitz, Katja
1 Institut für Mikrostrukturtechnik (IMT), Karlsruher Institut für Technologie (KIT)
2 Institut für Programmstrukturen und Datenorganisation (IPD), Karlsruher Institut für Technologie (KIT)
3 Institut für Funktionelle Grenzflächen (IFG), Karlsruher Institut für Technologie (KIT)

Abstract:

The split-synthesis approach offers a quick and easy method for producing a large diversity of substances for the discovery of novel protein ligands. Screening of the resulting one-bead-one-compound (OBOC) libraries with fluorescently labelled proteins is not trivial, however, since the resin beads can display significant autofluorescence. Here we present a simple two-channel microscopy image-based screening method for OBOC libraries on TentaGel MB-HMBA resin using tracers labelled with fluorophores. Bead images are taken at short exposure times in the RHO and FITC channels to keep photobleaching of the fluorophores at a minimum. Simple RGB colour vector addition ensures the identification of beads with high fluorescence intensities in the RHO channel. Pre-sorting of the library to exclude highly fluorescent beads is not necessary. The presented method is especially suited for small laboratories without automation equipment. By screening a library with a maximum of 117 649 peptoid hexamers we discovered 18 sequences that bind the human chemokine interleukin-8 (CXCL8) with affinities between 11 μM and 112 μM.


Verlagsausgabe §
DOI: 10.5445/IR/1000055939
Veröffentlicht am 12.01.2022
Originalveröffentlichung
DOI: 10.1039/C5AY02981C
Scopus
Zitationen: 2
Web of Science
Zitationen: 3
Dimensions
Zitationen: 2
Cover der Publikation
Zugehörige Institution(en) am KIT Institut für Funktionelle Grenzflächen (IFG)
Institut für Mikrostrukturtechnik (IMT)
Institut für Programmstrukturen und Datenorganisation (IPD)
Publikationstyp Zeitschriftenaufsatz
Publikationsjahr 2016
Sprache Englisch
Identifikator ISSN: 1759-9660, 1759-9679
KITopen-ID: 1000055939
HGF-Programm 47.02.06 (POF III, LK 01) Zellpopul.auf Biofunk.Oberflächen IFG
Erschienen in Analytical methods
Verlag Royal Society of Chemistry (RSC)
Band 8
Heft 20
Seiten 4142-4152
Vorab online veröffentlicht am 29.04.2016
Nachgewiesen in Dimensions
Scopus
Web of Science
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